This proposal examines the role of specific membrane proteolipids (Prs) in the calcification of the oral microorganism, Corynebacterium (Formerly Bacterionema) matruchotii. The experiments are based on the hypothesis that the presence of specific membrane proteins promote formation of phospholipid:Ca:Pi complexes (CPLX), subsequent hydroxyapatite (HA) deposition, and serve a structural role in the formation of ion channels. Proteolipids which support HA formation have an Mr of 10,000 on SDS-PAGE and when recombined with other C. matruchotii proteolipids, ion transport is observed. The Specific Aims are: 1.) To characterize the Mr 1O,OOO Pr(s) required for RA formation in vitro. 2. To compare the immunoreactive determinants of C. matruchotii Prs with those of other bacteria and the ability to calcify. 3.) To isolate Pr gene(s) using antibody probes. 4.) To examine the regulation of membrane mediated Pr-dependent HA formation in g. matruchotii. While our primary interest in this proposal is in microbial calcification, the technique developed and the principles established are directly relevant to membrane-mediated proteolipid-dependent calcification in vertebrate during endochondral ossification, dentinogenesis, or calcific disease. Biochemistry, immunology, and molecular biology will be used as tools to characterize the chemical and functional properties of these proteins during HA formation. Calcification in vitro will be used as a screening assay to identify and purify the 10K Pr that promotes HA deposition. Antibodies to this protein will be used to identify characteristics the promote HA formation via blocking experiments and by comparative analysis of immunoreactive determinants in CPLX. The antibody will also be used to clone the gene for this protein and to affinity purify the Pr. By varying culture physiology, the process of CPLX formation can be dissected, providing important new information applicable to septic stone a calculus formation as well as the basic process of calcification.